Nonalcoholic fatty liver disease (NAFLD) in humans has become an increasing public health problem paralleling the increase in obesity, type II diabetes, and changing dietary and activity patterns. The spectrum of NAFLD includes simple hepatic steatosis to steatohepatitis, fibrosis and cirrhosis. Published data and those from our preliminary studies suggest systemic insulin resistance and increased lipolytic flux are the major hallmarks of NAFLD in humans. An increased influx of fatty acids into the liver results in a higher rate of fatty acid oxidation, generation of a reactive oxygen species (ROS), increased glutathione consumption, and ultimately leads to mitochondrial dysfunction and propagation of disease. We hypothesize that hepatic insulin resistance and high fatty acid flux into the liver, in combination with increased oxidative stress, results in high methylation demands, a high rate of methionine transsulfuration, and glutathione synthesis. These changes in methionine metabolism may be exacerbated by altered folate metabolism as a result of gene polymorphism and by changes in hormones, cytokines and nutrient deficiency. We will examine these hypotheses by quantifying the rates of transmethylation and transsulfuration of methionine, and rate of glutathione synthesis in a well characterized population of hepatic steatosis and steatohepatitis patients using stable isotopic tracers. In addition, we will examine the responses to nutrient and fatty acid administration on the rate of glutathione synthesis. These data will be related to clinical and laboratory data, with measures of systemic insulin resistance and with methylenetetrahydrofolate reductase polymorphism. The proposed hypotheses are based upon strong preliminary data and published data in the literature. These studies will provide a mechanistic insight into the progression of NAFLD and will identify specific times in the disease when targeted interventions can be implemented and evaluated. PUBLIC HEALTH RELEVANCE: Nonalcoholic fatty liver disease has rapidly become the most common liver disease in the adult population worldwide. It has been recognized as the complication of obesity, type II diabetes and associated resistance to insulin action. The proposed studies are aimed at examining the metabolic mechanisms involved in the progression of this disease, so that specific targeted intervention strategies related to the stage of the disease can be identified and evaluated.